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Patients V. vuln i ficus septicemia Hepatic diseases, Immunocompromised conditions rapid progress and high mortality rate of >50%. V. vulnificus is confronted with dramatic environmental changes.
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Patients V. vulnificus septicemia Hepatic diseases, Immunocompromised conditions rapid progress and high mortality rate of >50% V. vulnificus is confronted with dramatic environmental changes V. vulnificus should establish coordinated spatiotemporal expression of various virulence genes in vivo Bacterial infection as assessed by in vivo gene expression. Proc. Natl. Acad. Sci. USA 1997 The authors’ previously reported 12 in vivo expressed genes by using in vivo-induced antigen technology Characterization and pathogenic significance of Vibrio vulnificus antigens preferentially expressed in septicemic patients. Infect. Immun. 2003
pyrHencodes UMP kinase Catalyzes phosphorylation of UMP to UDP It was reported that UMP kinase senses the environmental pyrimidine pool and directly regulates pyrimidine-specific CarP1 promoter of carbamoylphosphate synthetase of Escherichia coli responsible for the early stage de novo synthesis of pyrimidines. Limiting availability of pyrimidines in animal tissues might be responsible for the in vivo induction of the enzymes required for de novo biosynthesis of them
Construct pyrH gene-specific mutant strains significant decrease in virulence high frequency of curing Compromise enzymatic activity of PyrH by introducing site-directed mutations on the pyrH rather than by abolishing the gene
UMP binding amino acids RH DN The Arg62 was changed to His and the Asp77 to Asn
The introduction of point mutations at the essential amino acid residues induces a complete inhibition of PyrH activity.
The site-directed mutated gene was then cloned into the suicide vector pDM4, and the resulting plasmids were transferred into the V. vulnificus by conjugation. Effect of R62H/D77N mutation on the cytotoxicity and lethality of V. vulnificus. WT R62H/D77N MOI:100, 200 HeLa cell The release of cytosolic LDH from dead cells Complementation group
R62H/D77N strain has sufficiently attenuated virulence comparable to that of the pyrH insertional mutant and that PyrH enzyme activity has an important role in exerting full-blown virulence in vivo. 8-week-old specific-pathogen-free female CD-1 mice 900 µg of ferric ammonium citrate 30 min before bacterial challenge
Growth of the R62H/D77N in human ascites, HeLa cell lysates, and human serum. In vivo-like conditions The pyrH mutant should have defects in utilizing host-derived factors for their growth.
The pyrH Gene of Vibrio vulnificus Is an Essential In Vivo Survival Factor